Actions of phorbol ester tumor promoters are intimately related to modulation of queuine levels in the anticodons of specific transfer RNAs. Phorbol esters inhibit queuine uptake by cultured human skin cells, and they may also down-regulate expression and/or activity of the tRNA-modifying enzyme, tRNA-guanine ribosyltransferase. The resulting queuine-deficient tRNA is a common characteristic of neoplastic and undifferentiated cells. The ultimate goal of this research is to determine the significance of queuine-deficient tRNA in the processes leading to neoplasia (i.e. tumor promotion). The specific aims of this proposal are to characterize the control factors involved with phorbol ester inhibition of queuine uptake. This includes determination as to whether protein kinase C is directly involved with uptake inactivation, and to isolate and characterize a conditioned medium factor derived from early passage cultured human skin cells which potentiates phorbol ester inhibition of queuine uptake. Another aim is to define the potential phorbol ester induced down-regulation of tRNA-guanine ribosyltransferase activity and to purify the enzyme to homogeneity from human tissue sources. This will serve as a prelude to long term studies on the molecular biology of the enzyme and its expression, as a factor in tumor promotion through control of tRNA modification.